Immunotherapeutic methods and compositions employing antigens characteristic of malignant neoplasms

ABSTRACT

A method and composition for treating malignant neoplasia and acquired immune deficiency syndrome by once daily administration of a substance (such as human chorionic gonadotropin) characteristic of the tumor or acquired immune deficiency syndrome-afflicted cell in an amount less than the lowest amount necessary to provoke a humoral immune response, as exemplified by the existence of a negative wheal upon subcutaneous administration.

This is a division of application Ser. No. 692,822, filed Jan. 18, 1985,now U.S. Pat. No. 4,692,332, issued Sept. 8, 1987.

BACKGROUND OF THE INVENTION

The present invention pertains in general to immunotherapeutictechniques for alleviating the symptoms of malignant neoplasia and fortreating diseases of viral origin. In particular, the present inventionpertains to immunotherapeutic techniques useful in treatment of diseasestates such as feline leukemia, bovine leukemia and the acquired immunedeficiency syndrome (AIDS).

In order to protect the integrity of the organism, higher vertebratespossess an elaborate immune system which distinguishes foeignsubstances, which must provoke an immune response in order to beeliminated, from "self" substances, which are tolerated. The mechanismthat effectuates this discrimination between self and foreign substancesis known to involve interactions among types of white blood cells(leukocytes).

Upon exposure of the antigen to the circulating fluids of the body,substances capable of recognition by the immune system (antigens) comeinto contact with a type of leukocytes called a macrophage. Macrophagesare phagocytic cells and can therefore engulf and destroy materialswhich are not protected from them by size, surface texture (i.e.,smoothness), surface charge, or some other mechanism.

Once engulfed and processed by a macrophage, an antigen or a portionthereof is presented at the surface of the macrophage for contact withanother type of leukocyte called a thymocyte or T-cell. T-cells controlthe production of antibodies by yet another type of lymphocyte called aB-cell.

Antibodies are B-cell produced proteins which are capable of combiningwith an antigen in a reaction which is specific for that antigen. Anantibody only combines with certain portions (antigenic determinants) ofthe surface of the antigen, so that the antibody is specific to thedegree that the determinant with which it combines is not also found onother antigens.

The binding of an antibody to its corresponding antigen on the surfaceof a foreign cell has significant consequences related to thedestruction of that cell by the immune system. First, the coating of thecell by antibody facilitates ingestion of the cell by macrophages and byother types of phagocytes including killer (K) cells, which act todestroy antibody-coated cells but which do not require sensitization byprior exposure with macrophage-processed antigen, and polymorphonuclear(PMN) leukocytes. Second, the coating of a cell by antibody activates asystem of proteins, known as the complement system, in the liquid(plasma) fraction of the blood. Upon activation of this system,complement components also coat the foreign cell, which facilitatesphagocytosis. In addition, complement activation results in thestimulation of inflammatory cells, leading to production of chemicalswhich attract macrophages through a process called chemotaxis andleading to inflammatory hormone-like activation of cellular functions.Lastly, complement components act directly to break up (lyse) themembrane of the foreign cell. The portion of the immune responseinvolved with antigen-antibody and complement interactions is generallyreferred to as the humoral reaction.

T-cells, which regulates the humoral reaction, are of several types.These types of T-cells have been described as including helper (T_(H))cells, inducer (T_(I)) cells, regulator (T_(R)) cells, and suppressor(T_(S)) cells. [Herscowitz, Chapter 7 in Immunology III, Bellanti, J. W.Saunders, Philadelphia (1985)]. T_(I) and T_(H) cells are mobilized bycontact with processed antigen on the surface of macrophages. T_(H)cells are also activated by signals from T_(I) and from T_(R) cells.T_(R) cells are activated by signals from T_(I) and T_(S) cells.Mobilization of T_(S) cells occurs in response to signals from T_(R)cells or as a result of contact with antigen.

Introduction of an optimal amount of a foreign substance into the fluidsof the body initiates a process which results in production of antibodyby B-cells. In this process, B-cells respond to stimulation by T_(H)cells, which have in turn been stimulated by macrophages. Introductionof a persistent low level of some antigens or of a high level of anantigen results in a low-level of or in a lack of production of antibodydue to an interruption by T_(S) cells of the signals from T_(H) cells toB-cells. This interruption, called suppression, may be induced throughthe macrophage-T_(I) -T_(R) pathway or by direct stimulation of theT_(S) cells by antigen. Suppression of antibody production to a firstantigen may be overcome in a process known as contrasuppression throughthe stimulation of a subtype of T_(S) cells called contrasuppressorcells by a second antigen which is antigenically similar to the firstantigen. These contrasuppressor cells send a signal to T_(R) cells whichrender the T_(H) cells resistant to the activity of the suppressor T_(S)cells and which interrupt the suppressor signals of the suppressor T_(S)cells. See Gershon, et al., J.Exp.Med., 153: 1533-1546 (1981); Yamauchi,et al., J.Exp.Med., 153: 1547-1561 (1981); and Green, et al.,Ann.Rev.Immunol., 1: 439-463 (1983).

It is the balance of the actions of T_(H) helper and T_(S) suppressorcells which determines whether or not an immune response develops in thepresence of an antigen. Thus, as a practical matter, the functioning ofthe network of T-cells may be viewed in terms of the ratio of helper tosuppressor cells (T_(H) /T_(S)).

T-cells are also involved in another type of immune response, which issaid to involve cell-mediated immune (CMI) reactions. Contact of T_(H)cells with macrophage-processed antigen causes the T_(H) cells torelease interleukin II (IL-2), which activates cytotoxic (T_(CYT))T-cells and, in conjunction with gamma interferon also released by theT_(H) cells at this time, activates natural killer (NK) cells. BothT_(CYT) and NK cells kill foreign cells. T_(CYT) cells are particularlyinvolved with rejection and the destruction of tumor cells.

As is evident from the foregoing discussion, a general outline of thefunctioning of the immune system is available. However, many areas ofthe functioning of the immune system remain unclear. One of these areasrelates to the inability of the immune system to recognize certaincancers (malignant neoplasms) and cells infected with certain viruses[e.g., feline leukemia virus; bovine leukemia virus; and humanT-leukemia-lymphoma virus (HTLV), which is believed to be the causativeagent in AIDS].

In attempts to stimulate an immune response against a malignantneoplasm, many approaches have been aimed at the augmentation ofantitumor defenses by administrationo of adjuvants (immune enhancers orpotentiators). These approaches attempt to enhance nonspecificphagocytosis and killing of tumor cells by macrophages and T-cells. Suchappropriate employ infectious BCG mycobacteria, non-livingCorynebacterium parvum, glucan (a glucose polymer derived frommicroorganisms), or levamisole (an antihelminthic drug known to beuseful for stimulating CMI and the action of macrophages). Herberman, etal., Chapter 19 in Immunology III (Bellanti, ed.), W. B. Saunders Co.(1985), at page 343. The reported antitumor action of lysosome andpepsin lysates containing glycopeptides from the cell wall ofLactobacillus bulgarious [Bogdanov, et al., FEBS Letters, 57: 259(1975); Bogdanov, et al., Byulletin Eksperimental'noi Biologia iMeditsiiny, 84: 709 (1977)] and the treatment of malignant tumors withdestroyed Staphylococcus aureus [abstract of examined Japanese Pat.application No. 840486487] appear to fall in this category. Adjuvanttherapy has had varying degrees of questionable or limited success.Herberman, et al., supra.

The failure of the immune system to recognize malignant neoplasms isparticularly puzzling in view of the fact that certain characteristicsubstances (tumor markers) are present at levels which are elevatedabove normal in patients with various neoplastic disease states.Specifically, alphafetoprotein (AFP), carcinoembryonic antigen (CEA),and human chorionic gonadotropin (HCG) are oncofetal tumor markerswidely used in the investigation of patients with neoplasms of theliver, colon, and trophoblast, respectively. AFP has been found atlevels elevated above normal in fifty percent or more of patients withyolk sac tumors, hepatomas, retinoblastomas, embryonal carcinomas,breast carcinomas, and carcinomas of the uterine cervix, and has beenfound at elevated levels in between two and fifty percent of patientshaving carcinomas of the pancreas, melanomas, gastric carcinomas, basalcell carcinomas, bronchogenic carcinomas, leukemias, colon carcinomas,and nasopharyngeal carcinomas. CEA has been found at elevated levels infifty percent or more of patients having colon carcinomas,choriocarcinomas, pancreatic carcinomas, medullary thyroid carcinomas,familial medullary thyroid carcinomas, osteosarcomas, retinoblastomas,ovarian cystadenocarcinomas, mycosis fungoides, hepatomas, esophagealcarcinomas, adenocarcinomas of the uterine cervix, lung carcinomas,carcinomas of the small intestine, urinary bladder carcinomas, and renalcell carcinomas, and has been found at elevated levels in between nineand fifty percent of patients having neural crest tumors, breastcarcinomas, prostatic carcinomas, primary uveal carcinomas,neuroblastomas, fluids with malignancy, seminomas, basal cellcarcinomas, gastric carcinomas, laryngeal carcinomas, endometrialcarcinomas, uterine cervix intraepithelial carcinomas, carcinomas of thebuccal mucosa, craniopharyngiomas, embryonal rhabdomysarcomas,carcinomas of the orpharynx, brain tumors and testicular teratomas. HCGhas been found at elevated levels in the serum of fifty percent or moreof patients having choriocarcinomas, malignant interstitial cell tumorsof the testis, non seminomatous tumors of the testis, embryonalcarcinomas, and pancreatic carcinomas, and has been found at elevatedlevels in between six and fifty percent of the patients havingteratomas, ovarian adenocarcinomas, uterine cervix carcinomas,endometrial carcinomas, seminomas, gastric carcinomas, urinary bladdercarcinomas, breast carcinomas, colorectal carcinomas, bronchogenicsquamous cell carcinomas, mellanomas, and multiple myelomas. Otheruniversal oncofetal tumor markers, including tissue polypeptide antigen(TPA), which is associated with cell proliferation and which is notspecific for any species, are also known. See, Klavins, Annals ofClinical and Laboratory Science, 13: 275-280 (1983).

With respect to HGC, a chorionic gonadotropin-like antigen has beenfound in bacteria isolated from the urine of cancer patients, asindicated in Acevedo, et al., Infection and Immunity, 31: 487-494(1981), but not in the same species of bacteria obtained from any othersource tested. Furthermore, rat mammary adenocarcinoma cells and rathepatoma cells have been found to synthesize chorionic gonadotropin-likematerial, although no such material was found in the sera of the animalsbearing these neoplasms, in Kellen, et al., Cancer, 49: 2300-2304(1982); and Kellen et al., Cancer Immunol.Immunother., 13: 2-4 (1982).In the papers of Kellen et al., and in U.S. Pat. No. 4,384,995, asubunit of HCG conjugated to tetanus toxoid is used to prophylacticallystimulate an immune response to chorionic gonadotropin-like substancesby repeated injection with the conjugated material before exposure totumor cells known to bear a chorionic gonadotropin-like antigen.

Among the differences between prophylactic treatment with HCG and theadjuvant therapy approach is that the induction of an immune responsefor prophylactic purposes requires repeated injections over a period oftime in order to initiate the development of at least one population ofidentical B-cells (a clone) producing a given antibody to a tumorantigen and for antibody to be produced by that clone. On the otherhand, adjuvant therapy may result in antibody production by an existingclone of B-cells and thus has anti-tumor effects which may beimmediately observed. Therapeutic treatment (i.e., treatment after amalignant neoplasm is present) with HCG conjugated with tetanus toxoidraises the possibility of an uncontrollable Herxheimer-type reaction.The herxheimer reaction appears after treatment of syphilis patientswith a substance that is toxic to the causative spirochete bacteria,which thereupon die in massive numbers, releasing potentially fataltoxic substances into the blood stream. By analogy, at some as-yetunpredictable point in the induction of an immune response to a tumorantigen, a massive die-off of cancer cells may result in the death ofthe patient.

A luteinizing hormone releasing the factor (LHRF), sometimes genericallyreferred to as gonadorelin, which causes luteinizing hormone, apituitary gonadotropin, to be released from the pituitary, has been usedfor treating various tumors in U.S. Pat. No. 4,002,738 and No.4,071,622. Gonadorelin has also been used in the treatment of benignprostatic hyperplasia, a type of non-malignant but excess prostaticgrowth, in U.S. Pat. No. 4,321,260. However, no indication is providedin these patents that direct application of any gonadotropin may affectdestruction of malignant neoplasms. In addition, release of LH from thepituitary is subject to a feedback control independent of theadministered gonadotropin, so that how much, if any, LH is released isnot determinable merely from knowledge of an administered dose.Moreover, LHRF in combination with other substances may act to increasechorionic gonadotropin secretion by direct action on a tumor cell,further compounding the uncertain effect of LHRF administration. Kellen,et al., AACR Abstracts, 23: 235 (Mar. 1982) (Abstract 928).

In fact, Simon, et al., J.M.C.I., 70: 839-845 (1983), indicate thatdosages of gonadotropic and steroid hormones stimulate the growth ofdifferentiated carcinomas. These hormones included humanfollicle-stimulating hormone (FSH), HCG, human luteinizing hormone (LH),and cortisol. Thus Simon, et al. appears to support the idea that directadministration of gonadotropic or steroid hormones has a proliferativeeffect on malignant neoplasms.

Evidence for the suppression of the immune response against antigens ofneoplastic cells is provided by Akiyama, et al., J.Immunol., 131:3085-3090 (1983), wherein responsiveness of a mixed culture oflymphocytes from cancer patients and healthy donors was suppressed bythe introduction into the system of tumor cells from the cancerpatients. This suggests that among the lymphocytes of the cancerpatients were T_(S) cells specific for tumor-derived cells, inasmuch asthe response of cultures containing only lymphocytes from healthy donorswas not so suppressed.

Furthermore, antigen-specific T_(S) cells have been isolated from amouse having a plasmacytoma, which cells inhibited the in vitroinduction of a cytotoxic T-cell response against the tumor. Kolsch,Scand.J.Immunol., 19: 387-393 (1984). According to Kolsch, T_(S) cellsmay be activated and may dominate T_(H) cells by high and low doses ofantigen, but a critical, intermediate antigen dose which activates T_(H)cells at the same time as it activates the T_(S) cells, permits T_(H)cells to dominate. Thus, Kolsch indicates that there may be an antigendose at which a delicate balance is reached where T_(H) cells areactivated but at which T_(S) cells dominate the immune response.

In Loblay, et al., Aust.J.Exp.Biol.Med.Sci., 62: 11-25 (1984), it isindicated that the suppression produced by T_(S) cells in animals whichhave been exposed to an antigen is enhanced by a subsequentadministration of a sufficiently large dose of antigen. Perhaps it isnot so surprising, therefore, that attempts to induce contrasuppressionhave been aimed at supplying contrasuppressor cells, or substancestherefrom, rather than by direct induction of contrasuppression. See,Green, "contrasuppression: Its Role in Immunoregulation", in ThePotential Role of T-Cells in Cancer Therapy, Fefer, et al., eds., RavenPress, New York (1982); and Green et al., Ann.Rev.Immunol, 1: 439-463(1983).

SUMMARY OF THE INVENTION

Accordingly, a method according to the present invention for alleviatingsymptoms of malignant neoplasia in a disease victim comprises the stepof administering to the disease victim a member selected from the groupconsisting of characteristic substances of diseased cells of themalignant neoplasia victim and effective fragments and effectivederivatives thereof, in an amount which is less than the lowest amountnecessary to provoke a humoral immune response, as exemplified by thedetermination of a positive wheal upon subcutaneous injection.Illustrative of such a method is the once daily administration of acomposition including HCG and a lysate of S.aureus.

A further method according to the present invention for alleviatingsymptoms of acquired immune deficiency syndrome in a disease victimcomprises the step of administering to the disease victim a memberselected from the group consisting of characteristic substances ofdiseased cells of the acquired immune deficiency syndrome victim andeffective fragments and effective derivatives thereof, in an amountwhich is less than the lowest amount necessary to provoke a humoralimmune response, as exemplified by the determination of a positive whealupon subcutaneous injection.

A composition according to the present invention for alleviatingsymptoms of malignant neoplasia in a disease victim comprises a memberselected from a group consisting of characteristic substances ofdiseased cells of the malignant neoplasia victim and effective fragmentsand effective derivatives thereof, in an amount which is less than thelowest amount of the substance necessary to induce a humoral immuneresponse. The composition according to the present invention alsocomprises an immune enhancer in an amount less than the lowest amount ofthe substance necessary to provoke a humoral immune response, asexemplified by the determination of a positive wheal upon subcutaneousinjection. Illustrative of such a composition are compositions includingHCG and a lysate of S. aureus.

A further composition according to the present invention for alleviatingsymptoms of acquired immune deficiency syndrome in a disease victimcomprises a member selected from the group consisting of characteristicsubstances of diseases cells for the acquired immune deficiency syndromevictim and effective fragments and effective derivatives thereof, in anamount which is less than the lowest amount of the substance necessaryto induce a humoral immune response. This composition also comprises animmune enhancer in an amount less than the lowest amount of thesubstance necessary to provoke a humoral immune response, as exemplifiedby the determination of a positive wheal upon subcutaneous injection.Illustrative of such a composition are compositions including HCG and alysate of S. aureus.

DESCRIPTION OF THE PREFERRED EMBODIMENT

In a method according to the present invention, a substancecharacteristic of malignant neoplasia or AIDS is respectivelyadministered to a victim of malignant neoplasia or AIDS in an amountwhich is believed to be less than the lowest amount necessary to provokea humoral immune response (that is, to begin production of antibody).This dosage is administered daily until disappearance of the symptoms ofthe disease, and may be administered longer without harm if so desired.

In order to identify a dose lower than that required to provoke ahumoral immune response, a wheal produced upon subcutaneous injection ofthe therapeutic material is evaluated according to the procedure setforth in Moore, Clinical Medicine, 81, 16-19 (1974), wherein suchevaluation is employed to identify a dosage of vaccine useful in theeradication of the symptoms of influenza. Upon subcutaneous injection, awheal may be determined to be positive ten minutes after injection asblanched, hard, raised and discoid (regular, sharply demarcated edges,as though a disc has been cemented to the skin). A negative wheal,indicative of a dose below that necessary to provoke an immune response,is so absorbed at the end of ten minutes that is softer and flatter thanat injection, may have an irregular or ragged edge, and has grown lessthan an average of two millimeters in diameter.

Although the preferred dosage of 2 International Units (IU) wasinitially determined by skin tests of the horse of Example 1, theExamples below indicate that successful therapy may often be achieved byadministering a dose at a level of 2 IU. A dose of 2 IU appeared to below enough for any animal or human tested, even without firstdetermining optimum dose by a skin test. However, it is believed thatdetermination of a proper dosage, as exemplified above or otherwise, maybe used by those skilled in the art to refine the method according tothe present invention.

To complement the activity of the chorionic gonadotropin, and tosimultaneously guard against a toxic reaction induced by a rapidsloughing of necrotic tissue analogous to a Herxheimer-type reaction, abacterial lysate was added to each treatment vial as a broad-spectrumstimulator of cell-mediated immunity. No species specificity of responsewas observed, to the extent that a human has responded to either ofequine chorionic gonadotropin (ECG) or HCG, and that other animals ofthe examples have also responded favorably to treatment with either ECGor HCG.

AIDS patients have been reported as exhibiting a marked reduction in theratio of T_(H) to T_(S) cells. See, Cohen, British Journal of HospitalMedicine, 31, 250-259 (1984). AIDS has been defined as a disease atleast moderately predictive of a defect in cell-mediated immunity,occuring in a person with no known cause for diminished resistance tothat disease by the Centers for Disease Control.

It has been reported that the virus thought to cause AIDS is a type ofhuman T-cell leukemia-lymphoma virus (HTLV) known as HTLV-III virus andthat this virus is related to the virus which causes feline and bovineleukemia. Franklin, Science News, 126, 261 (1984). Feline and bovineleukemia viruses are known to be antigenically similar. Morgan, et al.,J.Virol., 46: 177-186 (1983). Accordingly, the discovery by the presentinventor that a dose of HCG which is lower than that required to provokea humoral immune response may be effective in the alleviation of thesymptoms of feline leukemia and of bovine leukemia, indicates thepotential effectiveness of an analogous form of treatment in thealleviation of the symptoms of AIDS as well.

Furthermore, feline leukemia virus (FLV) and HTLV are both retroviruses(also known as Type C viruses, RNA tumor viruses, leukemia viruses).Manzani, et al., Surv.Immunol.Res., 1: 122-125 (1982). A retrovirus maybe transmitted as an infectious particle containing viral genes encodedin the form of ribonucleic acid (RNA). Within an infected cell this RNAis encoded into deoxyribonucleic acid (DNA) by a viral enzyme calledreverse transcriptase. The DNA-encoded viral genes are thereafterintegrated with and replicated, transcribed and translated along withthe DNA-encoded genetic material of the infected cell. Lewin, Chapter 13in Genes, John Wiley and Sons, New York (1983). Such retrovirusesgenerally produce steady state infections where viral progeny arecontinually extruded by budding from the surfaces of host cells. In thisway, steady state viruses exhibit the clinical criteria for theinduction of tolerance in that there is a high dose inoculum of viralantigen, the virus-specific antigen persists, and tumor-specificantigens are developed and persist. See Herberman, et al., supra.

Therefore, the methods and compositions according to the presentinvention which are shown in the examples below to be effective in thetreatment of feline and bovine leukemia, among other neoplasticdiseases, are also expected to be effective in the treatment of AIDS.

Examples 1, 2 and 3 below relate to the treatment of horses afflictedwith two different types of malignant neoplasms.

Examples 4 and 5 below relate to the treatment of cats diagnosed ashaving feline leukemia.

Example 6 below relates to the treatment of cows afflicted with bovineleukemia.

Examples 7 and 8 below relate to the treatment of dogs afflicted withtwo different types of malignant neoplasm.

Example 9 below relates to the treatment of human patients afflictedwith malignant neoplasms.

Example 1

A horse with mastocytoma was treated with 2 IU per day of gonadotropinonly (i.e., without the bacterial lysate immune enhancer). Thegonadotropin used was equine chorionic gonadotropin, supplied by theW.A. Butler Company, or human chorionic gonadotropin, supplied by theAyerst Corp. (as A.P.L.-human chorioic gonatropin). In this example andin the examples which follow, a dosage of 2 IU per day of either equinechorioic gonatropin (ECG) or HCG was used.

The horse showed rapid and significant diminution of tumors beforesuccombing to an Herxheimer-type reaction. (No such adverse effect wasseen in other treated animals where an immune enhancer was used.)

Example 2

Three horses with melanoma were successfully treated to the point of thedisappearance of symptoms according to the procedure of Example 1 above,but with the addition of a bacterial lysate immune enhancer.

After the symptoms disappeared, the three animals received no furthertreatment but exhibited no recurrence of symptoms.

A suitable bacterial lysate for employment in this procedure is soldunder the name Staphage Lysate™, available from Delmont Labs. StaphateLysate™ is a bacteriologically sterile staphylococcal vaccine containingthe components of Staphylococcus aureus and culture medium ingredients(sodium chloride and ultrafiltered beef heart infusion broth). Thestaphylococcal components are prepared by lysing parent cultures of S.aureus, serologic types II and III, with a polyvalent staphylococcusbacteriophage. Each milliliter contains 120-180 million colony-formingunits of S. aureus and 100-1000 million staphylococcus bacteriophageplaque-forming units.

All treatment trials contained two units of gonadotropin plus 0.1 ccStaphage Lysate™ in each 0.5 cc shot.

Administration of Materials

Regardless of species, type of malignancy, or size of cancer, allanimals received a once-daily subcutaneous injection of the admixture ofchorionic gonadotropin and immune enhancer until the tumor or leukemiahad disappeared.

Tests and Results

All animals were diagnosed as having cancer by a licensed veterinarian.

EXAMPLE 3

One horse with an undifferentiated carcinoma of the face wassuccessfully treated according to the procedure of Example 2 to thepoint of the disappearance of symptoms. After disappearance of thesymptoms, the horse received no further treatment but exhibited norecurrence of symptoms.

EXAMPLE 4

A double-blind test of treatment of cats diagnosed as being infectedwith feline leukemia virus was conducted by a licensed testinglaboratory. Laboratory tests for feline leukemia were conducted by alicensed laboratory, and biopsy tissue from some cases was examined by aveterinary reference laboratory. In all cases animals were treated asabove, with termination of treatment coinciding with disappearance ofsymptoms. In no case was it necessary to resume therapy for a secondround of treatment. No side effects have been observed.

All animals receiving a placebo died within 7 days. Some treated animalswere still alive after six weeks, and were receiving no furthertreatment. Of especial interest was the observation that several treatedcats were not only asymptomatic, but also non-viremic. In any event,survival was prolonged and/or symptoms were alleviated for 6 (catsnumbered 2, 3, 11, 23 and 24) of 8 treated animals, as compared to theanimals receiving a placebo.

Cats numbered 4, 5, 16 and 22 received a placebo.

Cat number 4 died one day after the start of the test. Cat number 4 wasnot necropsied.

Cat number 5 showed no significant change in viremia. Tumor sizeremained constant. The cat was euthanized on day 10 of the test. Atnecropsy, the cat was observed to be moribund, thin, blind and anemic.Tumors were located on the tongue, eyes, pleural cavity, peririnal andperitoneal cavity.

Cat number 16 died six days after initial treatment. Upon necropsy,small metastatic tumors were observed in the lungs, and large tumorswere observed in the omentum.

Cat number 22 died six days after initial treatment. Upon necropsy,metastatic tumors were found in the lung, in the liver and in theomentum. The cause of death was renal hemorrhage into theretroperitoneal sublumbar region.

Cats numbered 2, 3, 7, 11, 15, 21, 23 and 24 received the experimentaltreatment.

In cat number 2, viremia decreased but the tumor remained the same size.Cat number 2 died 16 days after the start of the test. At necropsy, atumor was found which was open and which had drained. General lymph nodeenlargement, an enlarged liver, and a small tumor in the apex of theheart were also noted.

In cat number 3, viremia decreased to negative. The tumor decreased insize back to a normal condition. The cat was normal in appearance duringthe test.

Cat number 7 died four days after initial treatment. No necropsy wasperformed.

Cat number 11 exhibited a slight decrease in viremia. Tumor sizeremained the same. Cat number 11 died 19 days after initial treatment.Upon necropsy, tumors were found throughout the body cavity.

Cat number 15 showed no significant decrease in viremia. No change inthe tumor was observed. The cat died ten days after initial treatment.At necropsy, the cat was observed to be emaciated, and tumors were foundin the lungs, mediastinum, pericardia, pleura and illiac lymph nodes.

In cat number 21 the viremia was constant. The cat appeared normal. Uponnecropsy, an enlarged thymus and enlarged mesenteric lymph nodes wereobserved.

In cat number 23, viremia decreased through the course of the test. Catnumber 23 remained normal throughout the duration of the test.

In cat number 24, viremia remained constant. The cat appeared normalthroughout the duration of the test. Upon necropsy, an enlarged thymuswas observed, but the cat was otherwise normal.

EXAMPLE 5

Several dozen cats have been successfully treated according to theprocedure of Example 2 above by veterinarians in Ohio, Pennsylvania andNorth Carolina for feline leukemia. Based upon the reported observationsof the veterinarians, at least 80% and have returned to normal healthafter terminating therapy. For example, one cat had strength only tolift its chest wall to breathe, and its body weight was reduced from 13pounds to 6 pounds. After ten days of therapy, the cat was mobile andactive, and its weight had increased to ten pounds.

EXAMPLE 6

Twenty cases of bovine leukemia were treated according to the procedureof Example 2 above, with complete remission of symptoms for as long as13 months. Milk production of 17 cows was both restored and enhanced.Only three failures occurred, and in each case the cow had been moribundfor several days before initiation of therapy. Laboratory tests forbovine leukemia were conducted by a licensed laboratory, and biopsytissue from some cases was examined by a veterinary referencelaboratory.

EXAMPLE 7

A squamous cell carcinoma on the jaw of a six-month-old pup was treatedaccording to the procedure of Example 2 above. In spite of a "poorprognosis" from the veterinary reference laboratory examining the biposytissue, the dog entirely healed, as evidenced by sequential X-rayrecords. The dog has since been without treatment for nearly one year.

A squamous cell carcinoma on the shoulder of a 12-year-old dog wastreated according to the procedure of Example 2 above. After two daysthe dog could walk for extended periods, ate well, and the tumor becamewarm to the touch. Noticeable tumor shrinkage was observed after fivedays. After two weeks, the tumor was nearly resolved.

EXAMPLE 8

An anal tumor growing daily and laterally displacing the tail of a13-year-old dog was treated according to the procedure of Example 2above. The photographic record showed daily decrease in tumor sizebeginning on day three. The tumor resolved. The dog later died forunknown reasons. No necropsy was performed.

EXAMPLE 9

A limited number of human patients have been treated for cancersdiagnosed as terminal, including melanoma and cancers of the colon,breast, liver, pancreas and lung. The patients were treated according tothe procedure of Example 2 above, except that some patients received 0.2cc of immune enhancer per dose.

Of 9 patients treated at one location within a period of 21/2 years, alllived longer than the expected survival time indicated by their treatingphysicians, and only one died. The patient who died had discontinuedtreatment according to the present invention two months before death. Ofthe 8 surviving patients, 6 have survived 11/2 years or more sincebeginning treatment.

All 9 patients practiced the method according to the present inventionas a last resort, and all but one patient who died had had some form ofradiation or chemotherapy prior to beginning therapy according to thepresent invention. All 9 patients had diagnosed metastases indicative ofan advanced disease state. No side effects were observed.

It is not clear, however, how regularly all patients administered thecomposition according to the present invention. For example, somesurviving patients have decreased treatment to once or twice per weekwithout reduction in overall wellbeing. None of these patients (exceptfor the one patient who died) is known to have entirely discontinuedtreatment.

Sporadic treatment at other locations has not been as successful, sothat overall, about half of the treated patients are still surviving,while the other half have died. Due to the lack of autopsy data, it isnot clear whether or not all of the deaths of treated patients mayproperly be attributed to cancer or to other causes.

Patients who had not received maximum chemotherapy and radiationresponded most positively to the composition and method of the presentinvention. This suggests that the therapeutic agent of the presentinvention involves, at least in part, immune manipulation, and that thecells comprising the immune response in patients receiving traditionalanti-cancer therapy are compromised.

The effectiveness of treatment of malignant neoplasms, feline leukemia,and AIDS with gonadotropins may be explained in a number of ways. Therapidity of the response achieved according to the present invention,even with administration of HCG alone as in Example 3, suggests that thepresent invention is not operating merely by the initiation of a humoralimmune response. One explanation is that the low dosage of gonadotropinstimulates a contrasuppression reaction by tipping the balance of theT_(H) /T_(S) ratio in favor of activation of T_(H) cells, and that thisleads to the release of a pre-existing immune response (including a CMIresponse) against the disease. Another explanation may be that thegonadotropin acts in a negative feedback mechanism to turn offproduction of gonadotropin-like molecules by the malignant cells,leading to a reduction in gonadotropin-like cells on the surface of themalignant cells, leading in turn to a change in the surface charge fromthe negative charge associated with gonadotropin-like molecules to amore positive charge which facilitates ingestion by macrophages, orleading to exposure of otherwise hidden tumor antigens. However, it isnot intended that the present invention be limited to any explanation.

While the present invention has been described in terms of specificmethods and compositions, it is understood that variations andmodifications will occur to those skilled in the art upon considerationof the present invention.

For example, it is envisioned that various derivatives and fragments ofthe recited human chorionic gonadotropin and other tumor-orviral-specific antigens will be effective according to the presentinvention. In addition, although the preferred route of administrationis by subcutaneous injection, it is not intended to precludeintramuscular, intraperitoneal, or intravenous injection, intranasaladministration, or any other effective route of administration frombeing included within the scope of the present invention.

Also, inasmuch as other tumor markers, such as carcinoembryonic antigen,alphafetoprotein and tissue polypeptide antigen, are classified with HCGin relation to appearance in association with malignant neoplasms andare, therefore, likely to be similarly effective, it is intended thatthese substances be included within the scope of the present inventionas well.

Furthermore, human chorionic gonadotropin, follicle-stimulating hormone,luteinizing hormone and thyroid-stimulating hormone are eachglycoproteins composed of an α and a β subunit. The α subunit of HCGdiffers only slightly from an α subunit which is identical in each ofFSH, LH and TSH. Although the significance of the subunit structure isyet to be determined, both α- and β-HCG have been associated withtumors. See Acevedo, et al., Infection and Immunity, 31: 487-494 (1981).Accordingly, it is intended that these pituitary hormones be includedwithin the scope of the present invention.

It is contemplated that effective fragments and effective derivatives ofall of the foregoing suggested substances will be manufactured by thoseskilled in the art and employed according to the present invention.These fragments and derivatives are also intended to come within thescope of the invention as claimed.

Accordingly, it is intended in the appended claims to cover all suchequivalent variations which come within the scope of the invention asclaimed.

What is claimed is:
 1. A method for alleviating symptoms of felineleukemia or tumors in a diseased feline comprising the stepsof:administering to the diseased feline a member selected from the groupconsisting of human chorionic gonadotropin and equine chorionicgonadotropin and effective fragments and effective derivatives thereof,in an amount which is less than the lowest amount necessary to provoke ahumoral immune response, as exemplified by the presence of a positivewheal upon subcutaneous administration; and co-administering a lysate ofStaphylococcus aureus in an amount which is less than the lowest amountnecessary to provoke a humoral immune response in combination with themember, as exemplified by the presence of a positive wheal uponsubcutaneous administration.
 2. The method as recited in claim 1 whereinsaid administering step comprises the step of applying about 2 IU ofhuman chorionic gonadotropin or equine chorionic gonadotropin to adiseased feline. 29
 3. A composition for alleviating symptoms of felineleukemia or tumors in a diseased feline comprising:a member selectedfrom the group consisting of human chorionic gonadotropin and equinechorionic gonadotropin and effective fragments and effective derivativesthereof, in an amount which is effective for alleviating symptoms offeline leukemia or tumors in a diseased feline and which is less thanthe lowest amount of the substance necessary to provoke a humoral immuneresponse, as exemplified by the presence of a positive wheal uponsubcutaneous administration; and lysate of Staphylococcus aureus in anamount less than the lowest amount of the substance necessary to inducea humoral immune response in combination with said member as exemplifiedby the presence of a positive wheal upon subcutaneous administration. 4.The composition as recited in claim 3 wherein said amount of humanchorioic gonadotropin or equine chorionic gonadotropin if about 2 IU.